Publications by authors named "Joachim Lundahl"

59 Publications

B cell and monocyte phenotyping: A quick asset to investigate the immune status in patients with IgA nephropathy.

PLoS One 2021 19;16(3):e0248056. Epub 2021 Mar 19.

Karolinska Institutet, Clinical Science and Education, Södersjukhuset, Stockholm, Sweden.

Background: IgA nephropathy (IgAN) advances from multiple pathogenic "hits" resulting in poorly O-galactosylated IgA1 glycoforms (Gd-IgA1), production of antibodies and glomerular deposition of immune complexes. A sequence of immune responses arising from plasma cells, T cells and antigen presenting cells (APCs), causes glomerular injury. This study was designed to phenotype subsets of B cells, monocytes and T cells in the peripheral circulation and their association with inflammatory cytokines and kidney function in patients with IgAN, healthy controls (HC) and disease controls with autosomal dominant polycystic kidney disease (ADPKD).

Methods: Patients with IgAN (n = 13), median estimated glomerular filtration rate (eGFR) of 57 ml/min/1.73m2 (IQR 42-84), patients with ADPKD (n = 13) matched for kidney function, gender and age and gender and age-matched HC (n = 13) were recruited. CD3+ and CD3- peripheral blood mononuclear cells were isolated and profiled based on their specific surface markers for different subsets of monocytes, B and T cells and analyzed by flow cytometry. Cytokines were analyzed by ELISA.

Results: We observed a significant decrease in the proportion of pre-switched B cells and plasmablasts, but an increase in long-lived plasma cells in the peripheral circulation of IgAN patients compared to HC. The proportion of non-classical monocytes was significantly higher in IgAN patients compared to both HC and ADPKD. We also report an association between sCD40L levels and the proportion of pre-switched B cells, as well as sCD40L and MCP-1 levels and albuminuria in IgAN patients.

Conclusions: We applied an easy-access method to analyze subsets of immune cells as well as relevant inflammatory mediators in IgAN patients. Our data demonstrate an altered B cell profile that indicates a pathophysiological role of the B cell lineage and an increased proportion of non-classical monocytes that suggests their role in the disease process.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0248056PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7978284PMC
March 2021

Peanuts in the air - clinical and experimental studies.

Clin Exp Allergy 2021 04 21;51(4):585-593. Epub 2021 Feb 21.

Sachs' Children and Youth Hospital, Södersjukhuset, Stockholm, Sweden.

Background: Allergic reactions to food allergens usually occur after ingestion. However, fear of reactions to airborne peanut is a common concern for people with peanut allergy. There are no scientific reports on severe reactions with airborne peanut allergen.

Objective: To investigate the occurrence of allergic reactions in peanut-allergic children undergoing airborne peanut challenge and to determine levels of airborne peanut protein in a separate experimental evaluation.

Methods: Eighty-four children with peanut allergy underwent an airborne peanut challenge, 0.5 m from a bowl of peanuts for 30 min under controlled conditions. In a separate experiment, airborne peanut proteins from roasted and dry-roasted peanuts were collected at varying distances and at varying times with an electret SensAbues filter connected to an air pump. Collected airborne peanut proteins were extracted, dissolved and detected by ELISA. Basophil activation test was used to confirm biological activity.

Results: No moderate/severe allergic reactions to airborne peanut allergens were observed. Two children (2%) had mild rhino-conjunctivitis which required no treatment. The IgE-antibodies to peanut or Ara h 2 did not predict a reaction. In the experimental set-up, biological active peanut proteins were detected, in a very low amount, in median 166 ng/ml for dry-roasted and 33 ng/ml for roasted peanuts and decreased dramatically when the collection occurred at a greater distance (0.5-2 m) from the peanut source. Increased exposure time did affect the amount of collected peanut protein at 0 m, and the highest median was obtained after 60 min (p = .012); for time trend p = .0006.

Conclusions And Clinical Relevance: Allergic reactions to airborne peanut proteins are rare and cannot be predicted by high levels of IgE-antibodies to peanut or Ara h 2. Only small amounts of biologically active peanut proteins were detected in the air and seem unlikely to trigger moderate/severe allergic reactions.
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http://dx.doi.org/10.1111/cea.13848DOI Listing
April 2021

Adhesion molecule cross-linking and cytokine exposure modulate IgE- and non-IgE-dependent basophil activation.

Immunology 2021 01 29;162(1):92-104. Epub 2020 Oct 29.

Department of Clinical Science and Education, Karolinska Institutet, Södersjukhuset, Stockholm, Sweden.

Basophils are known for their role in allergic inflammation, which makes them suitable targets in allergy diagnostics such as the basophil activation test (BAT) and the microfluidic immunoaffinity basophil activation test (miBAT). Beside their role in allergy, basophils have an immune modulatory role in both innate immunity and adaptive immunity. To accomplish this mission, basophils depend on the capability to migrate from blood to extravascular tissues, which includes interactions with endothelial cells, extracellular matrix and soluble mediators. Their receptor repertoire is well known, but less is known how these receptor-ligand interactions impact the degranulation process and the responsiveness to subsequent activation. As the consequences of these interactions are crucial to fully appreciate the role of basophils in immune modulation and to enable optimization of the miBAT, we explored how basophil activation status is regulated by cytokines and cross-linking of adhesion molecules. The expression of adhesion molecules and activation markers on basophils from healthy blood donors was analysed by flow cytometry. Cross-linking of CD203c, CD62L, CD11b and CD49d induced a significant upregulation of CD63 and CD203c. To mimic in vivo conditions, valid also for miBAT, CD62L and CD49d were cross-linked followed by IgE-dependent activation (anti-IgE), which caused a reduced CD63 expression compared with anti-IgE activation only. IL-3 and IL-33 priming caused increased CD63 expression after IgE-independent activation (fMLP). Together, our data suggest that mechanisms operational both in the microfluidic chip and in vivo during basophil adhesion may impact basophil anaphylactic and piecemeal degranulation procedures and hence their immune regulatory function.
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http://dx.doi.org/10.1111/imm.13268DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7730031PMC
January 2021

A novel tool for clinical diagnosis of allergy operating a microfluidic immunoaffinity basophil activation test technique.

Clin Immunol 2019 12 24;209:108268. Epub 2019 Oct 24.

Department of Clinical Science and Education, Karolinska Institutet, Stockholm, Sweden; Sachs´ Children and Youth Hospital, Södersjukhuset, Stockholm, Sweden. Electronic address:

The Basophil Activation Test (BAT) is a valuable allergy diagnostic tool but is time-consuming and requires skilled personnel and cumbersome processing, which has limited its clinical use. We therefore investigated if a microfluidic immunoaffinity BAT (miBAT) technique can be a reliable diagnostic method. Blood was collected from allergic patients and healthy controls. Basophils were challenged with negative control, positive control (anti-FcεRI), and two concentrations of a relevant and non-relevant allergen. CD203c and CD63 expression was detected by fluorescent microscopy and flow cytometry. In basophils from allergic patients the CD63% was significantly higher after allergen activation as compared to the negative control (p<.0001-p=.0004). Activation with non-relevant allergen showed equivalent CD63% expression as the negative control. Further, the miBAT data were comparable to flow cytometry. Our results demonstrate the capacity of the miBAT technology to measure different degrees of basophil allergen activation by quantifying the CD63% expression on captured basophils.
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http://dx.doi.org/10.1016/j.clim.2019.108268DOI Listing
December 2019

Microfluidic Immunoaffinity Basophil Activation Test for Point-of-Care Allergy Diagnosis.

J Appl Lab Med 2019 09 9;4(2):152-163. Epub 2019 May 9.

Division of Nanobiotechnology, Department of Protein Science, Science for Life Laboratory, School of Engineering Sciences in Chemistry, Biotechnology and Health, KTH Royal Institute of Technology, Stockholm, Sweden;

Background: The flow cytometry-based basophil activation test (BAT) is used for the diagnosis of allergic response. However, flow cytometry is time-consuming, requiring skilled personnel and cumbersome processing, which has limited its use in the clinic. Here, we introduce a novel microfluidic-based immunoaffinity BAT (miBAT) method.

Methods: The microfluidic device, coated with anti-CD203c, was designed to capture basophils directly from whole blood. The captured basophils are activated by anti-FcεRI antibody followed by optical detection of CD63 expression (degranulation marker). The device was first characterized using a basophil cell line followed by whole blood experiments. We evaluated the device with ex vivo stimulation of basophils in whole blood from healthy controls and patients with allergies and compared it with flow cytometry.

Results: The microfluidic device was capable of capturing basophils directly from whole blood followed by in vitro activation and quantification of CD63 expression. CD63 expression was significantly higher ( = 0.0002) in on-chip activated basophils compared with nonactivated cells. The difference in CD63 expression on anti-FcεRI-activated captured basophils in microfluidic chip was significantly higher ( = 0.03) in patients with allergies compared with healthy controls, and the results were comparable with flow cytometry analysis ( = 0.04). Furthermore, there was no significant difference of CD63% expression in anti-FcεRI-activated captured basophils in microfluidic chip compared with flow cytometry.

Conclusions: We report on the miBAT. This device is capable of isolating basophils directly from whole blood for on-chip activation and detection. The new miBAT method awaits validation in larger patient populations to assess performance in diagnosis and monitoring of patients with allergies at the point of care.
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http://dx.doi.org/10.1373/jalm.2018.026641DOI Listing
September 2019

Associations of Fibroblast Growth Factor 23 with Markers of Inflammation and Leukocyte Transmigration in Chronic Kidney Disease.

Nephron 2018 4;138(4):287-295. Epub 2018 Jan 4.

Immunology and Allergy Unit, Department of Medicine Solna, Karolinska Institutet, Stockholm, Sweden.

Background: Patients with chronic kidney disease (CKD) show elevated levels of inflammatory markers and have an increased risk of infections as well as cardiovascular morbidity. Recent studies have implied effects of fibroblast growth factor 23 (FGF23) on inflammation in CKD. We analyzed potential correlations between levels of FGF23 with pro-inflammatory chemokines and markers of leukocyte transmigration in CKD patients.

Methods: One hundred three patients with CKD 2-5ND and 54 healthy controls, had biochemical markers in blood and urine analyzed according to routine protocol. Pro-inflammatory cytokines were analyzed by Milliplex technique and leukocyte CD11b adhesion molecule expression was measured by flow cytometry. FGF23 levels were measured with ELISA technique. Treatment of leukocytes from healthy blood donors with FGF23 was performed in vitro and effects analyzed by flow cytometry.

Results: Tumor necrosis factor-alpha, RANTES and interleukin (IL)-12 levels were significantly higher (p = 0.001, p < 0.001, and p < 0.001) in patients with CKD. Elevated FGF23 levels in the CKD group correlated to glomerular filtration rate, parathyroid hormone, urinary albumin excretion and phosphate as well as to IL-12 and RANTES. CD11b expression on resting granulocytes and monocytes, and on activated monocytes, was associated with FGF23. In vitro treatment of leukocytes with FGF23 reduced CD11b expression in resting as well as in formyl-methyinoyl-leucyl-phenylalanine-stimulated granulocytes (p = 0.03).

Conclusion: FGF23 levels are associated with various inflammatory markers such as pro-inflammatory cytokines and adhesion molecules on innate immune cells. However, further studies are warranted to define the direct role of FGF23 in modulation of the innate immune system in CKD.
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http://dx.doi.org/10.1159/000485472DOI Listing
September 2019

Altered basophil function in patients with chronic kidney disease on hemodialysis
.

Clin Nephrol 2017 Aug;88(8):86-96

Aims: Chronic kidney disease (CKD) leads to impairment of immune cell function. Given the potential role of basophils in the pathogenesis of CKD, we aimed to study the basophil responsiveness towards microbial antigen exposure, judged as adhesion molecule expression and degranulation, in CKD patients on hemodialysis.

Materials And Methods: We selected markers linked to two crucial biological phases: the transmigration and degranulation processes, respectively. For the transmigration process, we selected the adhesion molecules CD11b, active CD11b epitope, and CD62L and for the degranulation process CD203c (piecemeal degranulation marker), CD63 (degranulation marker), and CD300a (inhibitory marker of degranulation). We measured basophil responsiveness after stimulation of different activation pathways in basophils using lipopolysaccharide (LPS), peptidoglycan (PGN), formyl-methyinoyl-leucyl-phenylalanine (fMLP), and anti-FcεRI-ab.

Results: The expression of CD63 in basophils following activation by fMLP was significantly higher in the patient group compared to matched healthy controls, but no differences were observed after activation by anti-FcɛI. CD300a expression was significantly higher in patients following activation by fMLP and anti-FcɛI, and the active epitope CD11b expression was significantly higher in patients after LPS activation. In addition, we found that CD62L was not shed from the cell surface after activation with LPS and fMLP. A slight downregulation was noted after activation with anti-FcɛI in healthy controls.

Conclusion: Together, these data demonstrate that basophil functions related to adhesion and degranulation are altered in CKD patients on hemodialysis, which indicates a potential role for the basophil in the pathogenesis of complications related to infections.
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http://dx.doi.org/10.5414/CN108992DOI Listing
August 2017

Vitamin D receptor activation reduces inflammatory cytokines and plasma MicroRNAs in moderate chronic kidney disease - a randomized trial.

BMC Nephrol 2017 May 16;18(1):161. Epub 2017 May 16.

Department of Clinical Sciences, Karolinska Institutet, Danderyd University Hospital, Stockholm, Sweden.

Background: Chronic kidney disease (CKD) is a major risk factor for cardiovascular disease (CVD), partly due to endothelial dysfunction and chronic inflammation. Vitamin D treatment in end stage renal disease is suggested to modulate the immune system and lead to improved outcomes. We and others have demonstrated that treatment with vitamin D or activated vitamin D analogues protects the endothelial function in less severe renal disease as well. Since the endothelial protection might be mediated by vitamin D effects on inflammation, we assessed levels of pro-inflammatory cytokines and micro RNAs (miRs) in patients with moderate CKD, treated with an active vitamin D analogue (paricalcitol).

Methods: Thirty-six patients with moderate CKD were randomized to 12 weeks treatment with placebo, 1 μg, or 2 μg paricalcitol daily. Cytokines were measured by Milliplex 26-plex. Total RNA was isolated from plasma and miRs were determined by quantitative reverse transcription PCR analysis.

Results: Selected pro-inflammatory cytokines decreased significantly following treatment, while no change was observed in the placebo group. The micro RNAs; miR 432-5p, miR 495-3p, and miR 576-5p were significantly downregulated in the active treated groups, compared to the placebo group.

Conclusion: Paricalcitol treatment for 12 weeks in patients with moderate CKD reduces cytokines and micro RNAs involved in atherosclerosis and inflammation. The potentially protective role of vitamin D receptor activation in the inflammatory processes regarding the long-term outcomes in CKD patients warrants further studies.

Trial Registration: SOLID study; NCT01204528 , April 27, 2010.
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http://dx.doi.org/10.1186/s12882-017-0576-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5434555PMC
May 2017

Hemodialysis Patients Display a Declined Proportion of Th2 and Regulatory T Cells in Parallel with a High Interferon-γ Profile.

Nephron 2017 6;136(3):254-260. Epub 2017 Apr 6.

Immunology and Allergy Unit, Department of Medicine Solna, Karolinska Institutet, and Karolinska University Hospital, Stockholm, Sweden.

Background: A high prevalence of cardiovascular diseases (CVDs) and infections in patients with chronic kidney disease (CKD) arises partly due to a high inflammatory state and aberrations in immune cells function. Following in vitro stimulation of leukocytes with different T-cell mitogens, we observed a lower level of interleukin (IL)-2 and IL-10 production in CKD patients. To gain more knowledge as to whether this is the result of an alteration in T-cell function, we investigated the T-cell subsets profile and cytokine production in hemodialysis patients.

Methods: CD4+ cells were isolated from whole blood of 10 hemodialysis patients and 10 age- and gender-matched healthy controls. Following in vitro stimulation with an antigen-independent T-cell mitogen, Th1, Th2, and regulatory T (Treg) cell subsets were analyzed by flow cytometry through the expression of specific transcription factors. The levels of cytokines, interferon (IFN)-γ, IL-4, and IL-10 were analyzed by enzyme-linked immunosorbent assay in the supernatants.

Results: The proportion of CD4+CD25+FOXP3+ (Treg) and CD4+GATA3+ (Th2) cells was significantly lower in patients compared to healthy controls, while the proportion of CD4+T-bet+ (Th1) cells was similar. Moreover, levels of IL-4 were significantly lower in supernatants from patients, while IFN-γ levels were higher. IL-10 levels did not differ compared to those of the healthy controls.

Conclusions: Our findings indicate a diminished anti-inflammatory Treg, and Th2 cell profile in hemodialysis patients, accompanied by a high pro-inflammatory IFN-γ profile. Since this profile is characterized in CVDs, we propose that an imbalance between the inflammatory and anti-inflammatory responses may contribute to the pathogenesis of CVD in advanced CKD.
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http://dx.doi.org/10.1159/000471814DOI Listing
May 2018

Early Changes in Monocyte Adhesion Molecule Expression and Tumor Necrosis Factor-α Levels in Chronic Kidney Disease - A 5-Year Prospective Study.

Am J Nephrol 2016 8;44(4):268-275. Epub 2016 Sep 8.

Department of Nephrology, Karolinska Institutet, and Karolinska University Hospital, Stockholm, Sweden.

Background: Despite the absence of clinical symptoms, patients with chronic kidney disease (CKD) exhibit elevated levels of pro-inflammatory markers. To investigate whether it is possible to detect inflammatory activity and altered monocyte function at an early stage of renal disease, we studied patients with CKD stages 2-3 over 5 years.

Methods: The expression of adhesion molecules on monocytes at resting state and after stimulation with formyl-methionyl-leucyl-phenylalanine (fMLP), as well as oxidative metabolism capacity was measured with flow cytometry in 108 CKD patients and healthy controls. Soluble markers of inflammation, such as cytokines, were analyzed using the Milliplex technique.

Results: Patients showed significantly lower CD11b expression after stimulation during the 3rd (p = 0.002) and the 5th year (p < 0.001), together with a lower oxidative burst in response to fMLP over time (p = 0.02). The expression of CD62L on resting monocytes was lower during the 3rd (p = 0.001) and the 5th (p = 0.001) year in patients. Levels of tumor necrosis factor-α and RANTES were significantly increased (p = 0.001, p = 0.006) and interleukin-12 levels were also higher in CKD patients during the 5th year (p = 0.007).

Conclusion: Monocytes in CKD stages 2-3 show emerging functional abrasions, with altered adhesion molecule expression and impaired fMLP response. These findings suggest that a transformation of monocyte function occurs at an early phase of renal impairment and may together with increased plasma levels of pro-inflammatory cytokines contribute to the higher vulnerability of CKD patients to comorbidities, such as infections and cardiovascular disease.
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http://dx.doi.org/10.1159/000449290DOI Listing
January 2018

Differential expression of viral agents in lymphoma tissues of patients with ABC diffuse large B-cell lymphoma from high and low endemic infectious disease regions.

Oncol Lett 2016 Oct 16;12(4):2782-2788. Epub 2016 Aug 16.

Department of Oncology and Pathology, Karolinska Institute, 171 77 Stockholm, Sweden.

Diffuse large B-cell lymphoma (DLBCL), the most common type of non-Hodgkin's lymphoma (NHL) in adults, accounts for approximately 30-40% of newly diagnosed lymphomas worldwide. Environmental factors, such as viruses and bacteria, may contribute to cancer development through chronic inflammation and the integration of oncogenes, and have previously been indicated in cervical cancer, hepatocellular carcinoma, gastric cancer and lymphoproliferative disorders. In the present study, the presence of microbial agents was analyzed in the lymphoma tissue of patients with activated B-cell like (ABC) DLBCL. The present study compared two groups of patients from geographically varied regions that possess a difference in the prevalence of viral and other microbial agents. The patient populations were from Sweden (a low endemic infectious disease region) and Egypt (a high endemic infectious disease region). A differential expression of several viruses in lymphoma tissues was noted when comparing Swedish and Egyptian patients. JC polyomavirus (JCV) was detected in Swedish and Egyptian patients and, uniquely, the complete hepatitis B virus (HBV) genome was detected only in Egyptian lymphoma patients. None of these viruses were detected in control lymph tissues from Sweden or Egypt. In total, 38% of the Egyptian patients were found to have HBV surface antigens (HBsAgs) in their serum; however, HBsAgs were not found in any of the Swedish patients. The percentage of serum HBsAgs in Egyptian patients with ABC DLBCL was significantly increased compared with the general Egyptian population (P<0.05). The present study may support a notion that viral agents, including JCV and HBV, may be involved in the tumorigenesis of DLBCL in regions of high infectious disease.
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http://dx.doi.org/10.3892/ol.2016.5012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5038175PMC
October 2016

Eighteen years experience of granulocyte donations-acceptable donor safety?

J Clin Apher 2015 Oct 26;30(5):265-72. Epub 2014 Dec 26.

Department of Clinical Immunology and Transfusion Medicine, Karolinska University Hospital and Karolinska Institutet, Stockholm, Sweden.

Background: Granulocyte transfusions are given to patients with life-threatening infections, refractory to treatment. The donors are stimulated with corticosteroids ± granulocyte colony stimulating factor (G-CSF). However, data regarding the donors' safety is sparse. The objective was therefore to evaluate short- and long-term adverse events (AE) in G-CSF stimulated donors.

Study Design And Methods: All consecutive granulocyte donors from 1994 to 2012 were identified through our registry. From the donation records, the number of aphereses, stimulation therapy, AE, blood values post donation, and recent status were evaluated.

Results: One hundred fifty-four volunteer donors were mobilized for 359 collections. Age at first granulocyte donation was 43 years (median; range 19-64 years). Follow-up was 60 months (median; range 0-229 months). The dose of G-CSF per collection was 3.8 ug/kg body weight (median; range 1.6-6.0 ug/kg). Sedimentation agent was HES. Short-term AE were mild. Blood values 4 weeks post donation with minor reductions/elevations mostly resolved in later donations. Fourteen donors were excluded from the registry due to hypertension (4), diabetes (2), atrial flutter (1), breast carcinoma (1), urethral carcinoma in situ (1), MGUS (1), thrombosis (1), anaphylaxis (1), primary biliary cirrhosis (1), and unknown (1). Three donors are deceased due to diabetes, acute myocardial infarction, and unknown cause. All excluded/deceased donors except one were excluded/died at least 6 months after first granulocyte donation.

Conclusion: No serious short-term AE were observed. Due to the variability of diagnoses among excluded/deceased donors, we propose that it is less likely that granulocyte donations have a causative impact on these donors' exclusion or death.
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http://dx.doi.org/10.1002/jca.21373DOI Listing
October 2015

Activation of basophils is a new and sensitive marker of biocompatibility in hemodialysis.

Artif Organs 2014 Nov 9;38(11):945-53. Epub 2014 Apr 9.

Unit of Clinical Immunology and Allergy, Department of Medicine, Karolinska University Hospital Solna, Karolinska Institute, Stockholm, Sweden; Division of Proteomics and Nanobiotechnology, School of Biotechnology, Royal Institute of Technology, Stockholm, Sweden.

The hemodialysis procedure involves contact between peripheral blood and the surface of dialyzer membranes, which may lead to alterations in the pathways of innate and adaptive immunity. We aimed to study the effect of blood-membrane interaction on human peripheral basophils and neutrophils in hemodialysis with high- and low-permeability polysulfone dialyzers. The surface expression of CD203c (basophil selection marker) and CD63 (activation marker) after activation by the bacterial peptide formyl-methionyl-leucyl-phenylalanine (fMLP) or anti-Fcε receptor I (FcεRI) antibody and the absolute number of basophils was investigated before and after hemodialysis with each of the dialyzers. Moreover, the expression on neutrophils of CD11b, the CD11b active epitope, and CD88 was analyzed in the same groups of individuals. The expression of CD63 in basophils following activation by fMLP was significantly higher in the patient group compared with that in healthy controls, but no differences were observed after activation by anti-FcεRI. During the hemodialysis procedure, the low-flux membrane induced up-regulation of CD63 expression on basophils, while passage through the high-flux membrane did not significantly alter the responsiveness. In addition, the absolute number of basophils was unchanged after hemodialysis with either of the dialyzers and compared with healthy controls. We found no significant differences in the expression of the neutrophil activation markers (CD11b, the active epitope of CD11b, and CD88) comparing the two different dialyzers before and after dialysis and healthy controls. Together, these findings suggest that alterations in basophil activity may be a useful marker of membrane bioincompatibility in hemodialysis.
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http://dx.doi.org/10.1111/aor.12297DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4257079PMC
November 2014

Analysis of donor and recipient ABO incompatibility and antibody-associated complications after allogeneic stem cell transplantation with reduced-intensity conditioning.

Biol Blood Marrow Transplant 2014 Feb 22;20(2):264-71. Epub 2013 Nov 22.

Therapeutic Immunology Unit, Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden; Department of Clinical Immunology and Transfusion Medicine, Karolinska University Hospital, Stockholm, Sweden; Centre for Allogeneic Stem Cell Transplantation, Karolinska University Hospital, Stockholm, Sweden.

Allogeneic hematopoietic stem cell transplantation (HSCT) can be performed across the ABO blood group barrier. The impact of ABO incompatibility on clinical outcome is controversial. A retrospective analysis of 310 patients who underwent HSCT with reduced-intensity conditioning between 1998 and 2011 was performed to investigate the frequency and clinical implications of anti-RBC antibodies in passenger lymphocyte syndrome (PLS) after minor ABO mismatch (mm), persistent or recurring recipient type ABO antibodies (PRABO) after major ABO mm HSCT, and autoimmune hemolytic anemia (AIHA). Transplantation characteristics and clinical outcome were analyzed by univariate and multivariate analysis for groups with or without anti-RBC antibodies. ABO blood group incompatibility did not affect clinical outcome despite an increased requirement of blood transfusion. Twelve patients with AIHA, 6 patients with PLS, and 12 patients with PRABO post-HSCT were identified. AIHA did not affect overall survival (OS) or transplant-related mortality (TRM), but patients with AIHA had a lower incidence of grades II to IV acute graft-versus-host disease (P = .05). OS in the PLS group was 0% compared with 61% in the whole group receiving minor ABO mm transplants (P < .001). Comparing PRABO patients with those receiving a major ABO mm HSCT, the OS was 17% versus 73% (P = .002) and TRM was 50% versus 21% (P = .03). At our center, PLS after minor ABO mm and PRABO antibodies after major ABO mm HSCT are significant risk factors for decreased OS and TRM. Our results suggest that occurrence of unexpected ABO antibodies after HSCT warrant a wider investigation individual to find the underlying cause.
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http://dx.doi.org/10.1016/j.bbmt.2013.11.011DOI Listing
February 2014

Hemolytic disease of the fetus and newborn owing to anti-U, successfully treated with repeated intrauterine transfusions.

Immunohematology 2013 ;29(2):51-4

MD (corresponding author), Transfusion Medicine C2:66.

Hemolytic disease of the fetus and newborn (HDFN) owing to anti-U has rarely been reported. U is part of the MNS system.M and N glycoproteins are located on glycophorin A (GPA); Sand s antigens are on glycophorin B (GPB). Individuals who lack GPB are S- and s- and also lack U. The U- phenotype occurs almost exclusively in the African population and has a very low frequency (0.25%). Anti-U is of immunoglobulin G class and can cause hemolytic transfusion reaction and HDFN. In this report we present the use of a noninvasive method to detect anemia in the fetus and the subsequent use of intrauterine transfusion(IUT) with blood of a very rare phenotype. For the first time, we used deglycerolized and 3-week-old red blood cell units for IUT without signs of adverse reactions and with the expected effect on the hemoglobin value. We conclude that this transfusion strategy could be applied safely.
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December 2013

Leukocyte proliferation and immune modulator production in patients with chronic kidney disease.

PLoS One 2013 9;8(8):e73141. Epub 2013 Aug 9.

Unit of Clinical Immunology and Allergy, Department of Medicine, Karolinska University Hospital Solna, Karolinska Institutet, Stockholm, Sweden.

Introduction: In Chronic Kidney Disease (CKD), immune cells are affected by uremic retention toxins. Given this effect, we analyzed lymphocyte proliferative response and immune modulators production following in vitro stimulation.

Methods: Whole blood was drawn from healthy controls, patients with eGFR <20 ml/min/1.73 m(2) (Pre-dialysis, CKD stages 4 and 5) and hemodialysis patients (stage 5D). Peripheral cells were incubated for six days with pokeweed mitogen, concanavalin A, Staphylococcus enterotoxin A or influenza A vaccine. Peripheral lymphocyte proliferation was then analyzed by the "Flow-cytometric Assay of Specific Cell-mediated Immune response in Activated whole blood" (FASCIA) method, and cytokine profile in the cell supernatants was analyzed by the Milliplex multi-array method.

Results: The absolute number of lymphoblasts in response to mitogenic stimulation and the number of cells in each CD4+ and CD8+ subpopulation were similar comparing the three groups, except for a single decline in number of lymphoblasts after stimulation with Staphylococcus enterotoxin A, comparing dialysis patients with healthy controls. Levels of interleukin (IL)-2 (p=0.026), -10 (p=0.019) and -15 (p=0.027) in the Staphylococcus enterotoxin A-stimulated supernatant were lower in hemodialysis patients compared to healthy controls. Levels of IL-15 (p=0.017) from pre-dialysis patients and levels of IL-5 (p=0.019) from hemodialysis patients in influenza A vaccine-stimulated supernatants were also lower compared to controls. In pokeweed mitogen-stimulated supernatant, IL-2 levels (p=0.013) were lower in hemodialysis patients compared to pre-dialysis patients. TNF-α, IL-10, IL-12, IL-15, IL-8, MCP-1, IP-10, IFN-α2, IL-1α and eotaxin levels were all significantly higher in plasma obtained from CKD patients.

Conclusion: Our results suggest that T-cells from CKD patients have similar proliferative response to stimulation compared with healthy individuals. Moreover, however the immune cells show inability to produce selected cytokines, most likely due to the uremic milieu or dialysis procedure.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0073141PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3739766PMC
March 2014

Activation of Wnt/β-catenin pathway in monocytes derived from chronic kidney disease patients.

PLoS One 2013 23;8(7):e68937. Epub 2013 Jul 23.

Department of Laboratory Medicine, Division of Experimental Cancer Medicine-Clinical Research Center, Karolinska Institutet, Stockholm, Sweden.

Patients with chronic kidney disease (CKD) have significantly increased morbidity and mortality resulting from infections and cardiovascular diseases. Since monocytes play an essential role in host immunity, this study was directed to explore the gene expression profile in order to identify differences in activated pathways in monocytes relevant to the pathophysiology of atherosclerosis and increased susceptibility to infections. Monocytes from CKD patients (stages 4 and 5, estimated GFR <20 ml/min/1.73 m(2)) and healthy donors were collected from peripheral blood. Microarray gene expression profile was performed and data were interpreted by GeneSpring software and by PANTHER tool. Western blot was done to validate the pathway members. The results demonstrated that 600 and 272 genes were differentially up- and down regulated respectively in the patient group. Pathways involved in the inflammatory response were highly expressed and the Wnt/β-catenin signaling pathway was the most significant pathway expressed in the patient group. Since this pathway has been attributed to a variety of inflammatory manifestations, the current findings may contribute to dysfunctional monocytes in CKD patients. Strategies to interfere with this pathway may improve host immunity and prevent cardiovascular complications in CKD patients.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0068937PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3720736PMC
March 2014

Expression of microRNA-1234 related signal transducer and activator of transcription 3 in patients with diffuse large B-cell lymphoma of activated B-cell like type from high and low infectious disease areas.

Leuk Lymphoma 2014 May 31;55(5):1158-65. Epub 2013 Aug 31.

Department of Oncology and Pathology, Karolinska Institutet , Stockholm , Sweden.

Abstract Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease, and infectious agents are suspected to be involved in the tumorigenesis of DLBCL. MicroRNAs (miRNAs) are non-coding RNAs modulating protein expression. We compared miRNA expression profiles in lymph node tissues of patients with DLBCL of the activated B-cell like (ABC) type from two geographical areas with different background exposures, Sweden and Egypt. We showed previously that DLBCL tissues of the ABC-type in Swedish patients had a higher expression of signal transducer and activator of transcription 3 (STAT3) compared to Egyptian patients. Here, we analyzed the involvement of miRNAs in STAT3 regulation. miR-1234 was significantly up-regulated in Egyptian patients with DLBCL compared to Swedish patients (p < 0.03). The miR-1234 expression level correlated inversely with the expression of STAT3. The Stat3 protein was down-regulated in cells transfected with miR-1234, suggesting that STAT3 might be a potential target for miR-1234. miR-1234 and STAT3 might be involved in the tumorigenesis of DLBCL of ABC type and possibly associated with environmental background exposure.
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http://dx.doi.org/10.3109/10428194.2013.824077DOI Listing
May 2014

Effect of plasma-to-RBC ratios in trauma patients: a cohort study with time-dependent data*.

Crit Care Med 2013 Aug;41(8):1905-14

Department of Medical Epidemiology and Biostatistics, Karolinska Institutet, Stockholm, Sweden.

Background: A widespread approach today is to transfuse bleeding trauma patients with RBC concentrates and plasma at a 1:1 ratio. This regime is supported by a range of observational studies showing lower mortality in bleeding patients receiving equal volumes of plasma and RBCs. The rationale for this practice is still unclear with several studies failing to show any survival benefits of increased plasma use, perhaps due to a failure to account for the timing of transfused units.

Objective: To study the association between plasma-to-RBC ratios and risk of death in trauma patients, using appropriate methods.

Design, Settings, And Participants: In a retrospective cohort study, we assembled data on 741 transfused trauma patients at a large trauma center. Measures of transfusion therapy were assessed entirely time dependently, and relative risk of death was compared between patients receiving low to high plasma-to-RBC ratio (< 0.85 vs > 0.85).

Measurements And Results: In the time-dependent analyses, we saw no significant association between a low plasma ratio and the risk of death. However, age more than 75 years, injury severity score greater than 33, Glasgow Coma Scale less than 8, and systolic blood pressure lower than 90 mm Hg were all significantly associated with increased risk of death. Conversely, when the analyses were conducted with conventional methods, a strong protective effect of high plasma ratios was seen.

Conclusions: The key finding in our study is the strikingly different results produced by time-dependent analyses and the conventional analyses when studying survival and plasma-to-RBC ratio, supporting recent claims that prior studies showing benefit of high plasma ratios might have suffered from survival bias. There is a great need for further studies on the subject to enable improvements in treatment of massively bleeding trauma patients.
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http://dx.doi.org/10.1097/CCM.0b013e31828a3214DOI Listing
August 2013

The role of programmed cell death ligand-1 (PD-L1/CD274) in the development of graft versus host disease.

PLoS One 2013 4;8(4):e60367. Epub 2013 Apr 4.

Department of Laboratory Medicine, Division of Clinical Research Center, Experimental Cancer Medicine, Karolinska Institutet, Stockholm, Sweden.

Programmed cell death ligand-1 (PD-L1/CD274) is an immunomodulatory molecule involved in cancer and complications of bone marrow transplantation, such as graft rejection and graft-versus-host disease. The present study was designed to assess the dynamic expression of this molecule after hematopoietic stem cell transplantation in relation to acute graft-versus-host disease. Female BALB/c mice were conditioned with busulfan and cyclophosphamide and transplanted with either syngeneic or allogeneic (male C57BL/6 mice) bone marrow and splenic cells. The expression of PD-L1 was evaluated at different time points employing qPCR, western blot and immunohistochemistry. Allogeneic- but not syngeneic-transplanted animals exhibited a marked up-regulation of PD-L1 expression in the muscle and kidney, but not the liver, at days 5 and 7 post transplantation. In mice transplanted with allogeneic bone marrow cells, the enhanced expression of PD-L1 was associated with high serum levels of IFNγ and TNFα at corresponding intervals. Our findings demonstrate that PD-L1 is differently induced and expressed after allogeneic transplantation than it is after syngeneic transplantation, and that it is in favor of target rather than non-target organs at the early stages of acute graft-versus-host disease. This is the first study to correlate the dynamics of PD-L1 at the gene-, protein- and activity levels with the early development of acute graft-versus-host disease. Our results suggest that the higher expression of PD-L1 in the muscle and kidney (non-target tissues) plays a protective role in skeletal muscle during acute graft-versus-host disease.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0060367PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3617218PMC
November 2013

Patients with activated B-cell like diffuse large B-cell lymphoma in high and low infectious disease areas have different inflammatory gene signatures.

Leuk Lymphoma 2013 May 8;54(5):996-1003. Epub 2012 Nov 8.

Department of Oncology and Pathology, Karolinska Institutet, Stockholm, Sweden.

Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease with an association with inflammation and viral infections. We hypothesize that environmental factors may be involved in the pathogenesis of DLBCL. In this study, we compared gene expression profiles of lymph node tissues from patients with DLBCL from two different geographical areas with diverse environmental exposures. Specimens from Egyptian and Swedish patients with DLBCL as well as controls were studied. Gene expression analysis using microarray and quantitative polymerase chain reaction demonstrated significantly higher expression of signal transducer and activator of transcription 3 (STAT3) in Swedish as compared to Egyptian patients and control materials from both countries. This was confirmed at protein level using confocal microscopy. The receptor tyrosine kinase ROR1, a "survival factor" for malignant cells, was overexpressed and significantly related to the STAT3 expression pattern. The difference in the expression of genes involved in inflammatory responses and in the tumorigenic process of DLBCL might relate to infectious agents and/or other environmental exposures.
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http://dx.doi.org/10.3109/10428194.2012.738365DOI Listing
May 2013

Soluble interleukin-2 receptor alfa predicts renal outcome in IgA nephropathy.

Nephrol Dial Transplant 2012 May 22;27(5):1916-23. Epub 2011 Sep 22.

Nephrology Unit, Department of Medicine, Karolinska University Hospital, Karolinska Institutet, Stockholm, Sweden.

Background: Both systemic and mucosal IgA production are controlled by T lymphocytes and infiltrating T lymphocytes are involved in the progression of interstitial fibrosis in chronic kidney disease (CKD). Since the concentration of soluble interleukin-2 receptor alfa (sIL-2Ra) reflects the degree of T cell activation over time, we studied the impact of interleukin-2 receptor alfa levels on disease progression in patients with biopsy-proven IgA nephropathy (IgAN), a disease in which 20-30% of the patients progress to end-stage renal failure.

Methods: sIL-2Ra plasma levels were measured in 194 patients (median age 39 years, 70% men) and 84 matched controls. One hundred and seventy-nine of the patients, with an estimated glomerular filtration rate (GFR) of ≥15 mL/min/1.73m(2) at baseline (CKD Stages 1-4), were followed for up to 15 years (median 52 months; range 12-188). sIL-2Ra was evaluated as a risk marker for severe renal progression, here defined by the development of CKD Stage 5 (GFR <15 mL/min/1.73m(2)), a 50% decline in GFR during the follow-up period or a 30% GFR decline within 5 years of follow-up. In 51 patients, upon whom a renal biopsy had been performed within 2 years of IL2-Ra measurement, the biopsies were scored according to the Oxford classification. The correlations between the histopathological findings and the sIL-2Ra levels were examined.

Results: sIL2-Ra levels were significantly higher in patients than in controls (P < 0.001). sIL-2Ra levels in the upper third tertile predicted a severe renal outcome, even after adjustment for the main clinical risk factors: time average albuminuria and GFR at baseline (Relative risk 5.35, P < 0.001). sIL-2Ra levels also correlated significantly to the yearly GFR slope (β = -0.24, P = 0.01). According to the Oxford classification, the presence of >25% tubular atrophy/interstitial fibrosis (T1-2) was associated with higher sIL-2Ra levels, after adjustment for serum creatinine levels, if analysed within 4 months [n = 24, odds ratio (OR) 1.0, P = 0.044] or within 2 years from the kidney biopsy (n = 51, OR 1.0, P = 0.017).

Conclusions: The plasma levels of sIL-2Ra were predictive of long-term renal disease progression in a large cohort of patients with biopsy-proven IgAN. Further studies are warranted to evaluate if sIL-2Ra levels can feasibly contribute in the monitoring of effects of treatment, aimed to prevent the progression of interstitial fibrosis and progressive glomerulosclerosis in IgAN.
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http://dx.doi.org/10.1093/ndt/gfr554DOI Listing
May 2012

Gene expression analysis using long-term preserved formalin-fixed and paraffin-embedded tissue of non-small cell lung cancer.

Int J Oncol 2011 Apr 8;38(4):1075-81. Epub 2011 Feb 8.

Department of Oncology and Pathology, Karolinska Institutet, Stockholm, Sweden.

This study was performed to evaluate RNA extraction and gene expression analysis of non-small cell lung cancer (NSCLC) using formalin-fixed and paraffin-embedded (FFPE) specimens stored for more than 20 years by quantitative PCR (qPCR) and DNA microarrays. Long-term preserved FFPE materials enable large retrospective studies correlating molecular features with therapeutic response and clinical outcome. qPCR was used to evaluate RNA extraction methods and to compare DNA microarray gene expression profiles of FFPE and fresh frozen (FF) tissue. The Ambion RecoverAll kit appeared to be suited for RNA extraction of long-term preserved FFPE tissues. Microarray analysis using the Affymetrix platform displayed a high degree of correlation for endogenous control genes comparing FF and FFPE tissues and identified known NSCLC signature genes in both specimens. We conclude that high quality gene expression signatures can be recognized using the Affymetrix gene expression platform on FFPE tissue stored for more than 20 years. However, a general interpretation must be done with caution as different FFPE procedures have varying effects on RNA quality.
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http://dx.doi.org/10.3892/ijo.2011.936DOI Listing
April 2011

Expression of neutrophil SOD2 is reduced after lipopolysaccharide stimulation: a potential cause of neutrophil dysfunction in chronic kidney disease.

Nephrol Dial Transplant 2011 Jul 2;26(7):2195-201. Epub 2010 Nov 2.

Department of Nephrology, Skåne University Hospital, Malmö, Sweden.

Background: Neutrophils from patients with chronic kidney disease (CKD) are dysfunctional and thus a contributing factor to the risk of infections. The mechanisms for leucocyte dysfunction in CKD are not fully understood. It is known that lipopolysaccharide (LPS) activates transcription of several genes encoding proinflammatory cytokines. We therefore aimed to study the effect of LPS on neutrophil expression of genes related to the inflammatory response to address the hypothesis that LPS-induced gene transcriptions are altered in CKD patients.

Methods: We analysed gene expression of LPS-stimulated neutrophils from 30 patients with CKD and 15 healthy controls. Superoxide dismutase-2 (SOD2), IL1A, IL-1R1, IL-1R2 and IL8RA gene expression from both neutrophils and differentiated HL60 cells were measured by quantitative polymerase chain reaction. Differentiated HL60 cells were stimulated with phorbol-12-myristate-7-acetate (PMA) after inhibition of SOD2 by small interfering RNA followed by respiratory burst assessment using flow cytometry.

Results: LPS stimulation induced a significant mobilization of CD11b on neutrophils from CKD and healthy controls. Upregulation of SOD2, IL1A, IL-1R1 and IL-1R2 gene expression in neutrophils from healthy controls after LPS stimulation was contrasted by no change in gene transcription (IL-1R1 and IL-1R2) or even a downregulation in patients with CKD (SOD2 and IL1A). Inhibition of SOD2 reduced the PMA-induced respiratory burst and IL1A, IL-1R1, IL-1R2 and IL8RA gene expression in neutrophil-differentiated HL60 cells.

Conclusions: Because of the critical role of SOD2 in the generation of hydrogen peroxide during phagocytosis, downregulation of SOD2 gene expression after LPS stimulation in neutrophils from patients with CKD indicates a potential mechanism for neutrophil dysfunction and cytokine dysregulation in these patients.
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http://dx.doi.org/10.1093/ndt/gfq673DOI Listing
July 2011

Inflammatory cytokines in saliva: early signs of metabolic disorders in chronic kidney disease. A controlled cross-sectional study.

Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2010 Nov;110(5):597-604

Department of Dental Medicine, Public Dental Service, Karolinska University Hospital, Karolinska Institutet, Solna, Sweden.

Objective: The aim of this study was to evaluate correlations between levels of cytokines in secreted stimulated saliva in patients with chronic kidney disease (CKD) and hyposalivation.

Study Design: Seventy patients with clearance <20 mL/min/1.73 m(2) were evaluated; 40 were predialysis, 21 hemodialysis, and 9 peritoneal dialysis, and they were matched with 70 control subjects. Salivary flow rate was measured and submandibular/sublingual saliva collected. Analyses were performed for whole protein content using a protein assay, and levels of tumor necrosis factor (TNF) α, interleukin (IL) 1β, γ-interferon (γ-INF), IL-6, IL-8, IL-10, monocyte chemotactic protein (MCP) 1, and soluble intercellular adhesion molecule (sICAM) 1, by using Luminex technology.

Results: Patients with CKD had lower (P = .03) stimulated salivary secretion rate and higher salivary whole protein concentration (P = .002) than control subjects. Concentrations of IL-8 (P = .03) and MCP-1 (P = .002) were decreased and TNF-α/IL-10 (P = .05) and IL-8/IL10 (P = .03) ratios were decreased in CKD patients. CKD patients with low secretion levels of stimulated saliva expressed decreased levels of TNF-α (P = .04), IL-1β (P = .02), γ-INF (P = .03), IL-6 (P = .003), IL-8 (P = .005), MCP-1 (P = .006), and sICAM-1 (P = .02).

Conclusions: Salivary cytokines and secretion rates are significantly decreased in CKD patients. Further research is necessary to understand operating mechanisms and clinical implications of the down-regulation of inflammatory markers in saliva.
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http://dx.doi.org/10.1016/j.tripleo.2010.07.007DOI Listing
November 2010

Safety and tolerability of a modified filter-type device for leukocytapheresis using ACD-A as anticoagulant in patients with mild to moderately active ulcerative colitis. Results of a pilot study.

J Clin Apher 2010 ;25(5):287-93

Unit of Clinical immunology, Department of Clinical Immunology and Transfusion Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden.

Cellsorba™ is a medical device for leukocytapheresis (LCAP) treatment of ulcerative colitis (UC). Cellsorba™ EX Global type has been developed from Cellsorba E for intended use with ACD-A as anticoagulant. We evaluated safety and efficacy of the modified Cellsorba using ACD-A in a pilot trial comprising patients with active UC, despite receiving 5-ASA. A total of 10 LCAP treatments/patients were administered. Safety assessment focused on clinical signs and symptoms, hematological variables, as well as levels of bradykinin and IL-6. Efficacy was determined using the Mayo clinical/endoscopic scoring index as well histological assessment of biopsies. Additional aim was to evaluate the impact of apheresis system lines and filter on selected regulatory molecules. All six subjects completed the trial without any serious adverse events. WBC, platelet counts, and levels of bradykinin and IL-6 were not significantly affected. The median Mayo score decreased from 8.0 to 3.5 at week 8 (and to 2 at week 16 for the responders). Four patients were responders, of whom two patients went into remission. Median histological scores decreased from 3.5 to 2.0 in these four patients. Concentration of LL-37 increased within the apheresis system lines. LCAP with Cellsorba EX using ACD-A as anticoagulant was found to be a safe and well-tolerated procedure in patients with active UC. The positive impact on efficacy parameters merits further evaluation in a controlled fashion.
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http://dx.doi.org/10.1002/jca.20255DOI Listing
February 2011

Neutrophil activation during transmigration in vivo and in vitro A translational study using the skin chamber model.

J Immunol Methods 2010 Sep 5;361(1-2):82-8. Epub 2010 Aug 5.

Department of Clinical Immunology, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden.

Neutrophil transmigration can be studied in vitro by use of the transwell model and in vivo by the skin chamber model. Activation during transmigration involves translocation of secretory vesicles and granules to the plasma- and phagolysosome membranes. In this study, we compared the skin chamber model with the transwell model, focusing on the mobilization of CR1 (CD35), CR3 (CD11b/CD18) and CD63 from intracellular vesicles and granules. In addition, functional responses towards a bacterial related stimulus, formyl-methionyl-leucyl-phenylalanine (fMLP), in terms of CR3 expression and production of reactive oxygen species (ROS) were assessed. Discrepancies between the skin chamber model and the transwell model were observed. The expression of CR1 increased following in vivo transmigration (p<0.001) and, in contrast, decreased following in vitro transmigration (p=0.004). Furthermore, CR1 was mobilized following an isolation procedure included in the transwell model. The expression of CR3 increased following both in vivo (p<0.001) and in vitro (p=0.03) transmigration. However, in vitro transmigration did not influence the fMLP induced CR3 expression which was significantly increased following in vivo transmigration (p=0.01). In addition, the fMLP induced production of ROS was significantly reduced following in vitro transmigration (p=0.002) but unaltered after in vivo transmigration, indicating differences between the impact of the two systems on cellular activation. The observed discrepancies between the two models might be partly explained by granule mobilization and neutrophil priming, induced during the isolation procedure included in the transwell model, which results in an altered cellular activation. Therefore, mobilization of granules needs to be accounted for when interpreting data from different model systems.
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http://dx.doi.org/10.1016/j.jim.2010.07.015DOI Listing
September 2010

Low immunogenicity allows Staphylococcus epidermidis to cause PD peritonitis.

Perit Dial Int 2011 Nov-Dec;31(6):672-8. Epub 2010 May 6.

Clinical Microbiology, Karolinska University Hospital and Karolinska Institutet, Stockholm, Sweden.

Background: Peritonitis is a common and serious complication of peritoneal dialysis (PD). Coagulase-negative staphylococci from the patient's own skin flora are the most commonly found micro-organisms.

Objective: In the present study we aim to elucidate the immune response in the early stage of infection and to clarify the importance of bacterial attachment to fibrinogen.

Methods: Clinical Staphylococcus epidermidis isolates collected from PD peritonitis or the residential skin flora of healthy individuals were used to infect monocytes, macrophages, and peripheral blood mononuclear cells (PBMC) in the presence or absence of fibrinogen. The S. epidermidis strain HB (fbe(+)), expressing the fibrinogen-binding protein Fbe, and its isogenic mutant ST056 (fbe(-)) were used to study the impact of Fbe during cell infection. Immune induction was measured as interleukin-8 (IL-8) production determined by ELISA. Modulation of CD11b/CD18 expression in neutrophils incubated in conditioned medium from these experiments was analyzed in order to judge the cellular response.

Results: S. epidermidis causing peritonitis was less immunogenic compared to strains belonging to the residential skin flora, as measured by IL-8 induction in monocytes and CD11b/CD18 expression in neutrophils. At low bacterial concentrations, attachment to fibrinogen was a prerequisite for an IL-8 induction in monocytes and PBMC. The fibrinogen-binding protein Fbe did not, however, influence immune induction under this condition.

Conclusions: We suggest that S. epidermidis strains may be able to cause clinical infection by evoking an inadequate immunological response in the early stage of infection. Bacterial attachment to fibrinogen is a relevant event during this phase but independent of the fibrinogen-binding protein Fbe.
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http://dx.doi.org/10.3747/pdi.2009.00150DOI Listing
April 2012

Infliximab in clinical routine: experience with Crohn's disease and biomarkers of inflammation over 5 years.

Eur J Gastroenterol Hepatol 2009 Oct;21(10):1168-76

Department of Medicine, Gastroenterology and Hepatology Unit, Karolinska Institutet, Stockholm, Sweden.

Introduction: Infliximab was launched for the treatment of Crohn's disease (CD) in 1999. We set up a follow-up protocol to meticulously study disease development with repeated infusions of infliximab.

Aim: To follow the effects of infliximab treatment on disease activity, blood chemistry, quality of life, plasma nitrite, and titers of Saccharomyces cerevisiae antibodies (ASCA).

Methods: During 1999-2008, CD patients were monitored for disease activity by Harvey-Bradshaw index, blood chemistry with hemoglobin, albumin, C-reactive protein, platelet count, leukocyte count and creatinine, quality of life by the Short Health Scale, and plasma nitrite. During the first year of treatment, follow-up was done repeatedly before and 1 week after each infusion and thereafter every year before the last infusion for 5 years. ASCA was analyzed by flow cytometry with fluorescein isothiocyanate-labelled antibodies.

Results: A total of 1061 infusions were given to 103 patients; 92 responders and 11 nonresponders. Responders were further monitored and Harvey-Bradshaw index decreased with infusions during the first year of treatment (P < 0.0001), whereas hemoglobin (P < 0.01) and albumin (P <0.001) increased, C-reactive protein (P < 0.01) decreased, platelets (P <0.001) increased, and leukocytes (P< 0.01) decreased. Creatinine was not affected. Short Health Scale (questions analyzed separately) decreased (P < 0.0001), and nitrite (P < 0.001) increased. During the next 4 years the improved values remained stable. Adverse effects were noted among 32% of the patients; local circulatory reactions being most common. No correlation between ASCA titers and inflammatory activity or infliximab treatment was found.

Conclusion: Infliximab treatment is highly effective in responders and maintains symptomatic improvement and low inflammatory activity over years in CD patients.
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http://dx.doi.org/10.1097/meg.0b013e32832b125cDOI Listing
October 2009

Staphylococcus epidermidis isolated from newborn infants express pilus-like structures and are inhibited by the cathelicidin-derived antimicrobial peptide LL37.

Pediatr Res 2009 Aug;66(2):174-8

Department of Woman and Child Health, Karolinska Institutet, Stockholm 17176, Sweden.

Coagulase-negative staphylococci and its subtype Staphylococcus epidermidis are major indigenous Gram-positive inhabitants of the human skin. Colonization occurs in direct connection with birth and terrestrial adaptation. This study focuses on factors that may influence skin colonization of the newborn infant that relates to the immune status of both the bacteria and the host. Skin is an effective barrier against bacteria, and this function is partly mediated by the presence of antimicrobial peptides including human cathelicidin peptide LL37. Gram-positive bacteria have been described to have adhesive pili on their surface that mediates specific attachment to the host. Here, we identify, by negative staining transmission electron microscopy (EM), two different types of pilus-like structures commonly expressed on S. epidermidis isolated from newborn infants. We also show that the cathelicidin antimicrobial peptide LL37, constitutively expressed in the skin barrier of the newborn, significantly inhibited growth of S. epidermidis indicating its importance for the ecological stability of the skin microbiota. Further studies are required to elucidate molecular mechanisms of host-microbe interactions, both for the maintenance of a mutually beneficial homeostatic relationship and for the protection of self when it results in overt disease.
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http://dx.doi.org/10.1203/PDR.0b013e3181a9d80cDOI Listing
August 2009
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